Dopaminergic (DA) neurons in the ventral tegmental area (VTA) provide the DA innervation of the nucleus accumbens;this mesolimbic DA pathway is important for the rewarding properties of ethanol. Ethanol directly excites acutely dissociated DA VTA neurons, in the absence of input from surrounding cells. Ethanol excitation of DA VTA neurons is associated with a reduction in the action potential afterhyperpolarization (AHP) suggesting that it is due to a decrease in a potassium (K) current which contributes to the AHP. Ethanol excitation is completely blocked by quinidine, but not by the other K channel blockers apamin, tetraethylammonium, barium or cesium. In whole cell voltage clamp experiments, ethanol reduced the sustained outward current evoked by depolarizing voltage steps from aholding potential of-40 mV (to inactivate A-current). These data suggest that ethanol excites DA VTA neurons by reducing a non- or slowly inactivating, quinidine-sensitive, K current of the delayed rectifier type. The specific aims of the present application are two-fold. 1) Electrophysiological and pharmacological characterization of this ethanol- sensitive K current in DA VTA neurons, and determination of the cutoff for ethanol excitation and for reduction of the ethanol-sensitive K current by longer chain length alcohols. 2) Molecular biological studies to try to identify the native ethanol-sensitive K current in terms of cloned K channels of known structure. Candidate channels have been selected according to their electrophysiological and pharmacological similarityto the native channel. RT-PCR on pooled neurons and single cell RT-PCR will be used to determine which candidate channel mRNAs are expressed in DA VTA neurons. Then immunohistochemistrywill be used to see which channel proteins are actually present on the soma and dendrites of DA VTA neurons. Finally, the most likely candidate channels will be expressed in Xenopus oocytes and the electrophysiological properties and cutoff for longer chain alcohols will be determined for these K currents and compared to the properties of the native ethanol-sensitive K current in DA VTA neurons. These data should give important information on the mechanism by which ethanol directly excites DA VTA reward neurons. Taken together, the electrophysiological and molecular biological data should help to identify the ethanol-sensitive native channel on DA VTA reward neurons and therefore point to a gene responsible for its expression. Such a discovery could have major implications for understanding genetic differences in ethanol effects on the mesolimbic reward pathway and how changes in the response of DA VTA neurons during chronic ethanol consumption leads to alcohol craving and addiction.